Molecular modeling of G-protein coupled receptor kinase 2: Docking and biochemical evaluation of inhibitors

Abstract

G-protein coupled receptor kinase 2 (GRK2) regulates the activity of many receptors. Because potent inhibitors of GRK2 are thus far limited to polyanionic compounds like heparin, we searched for new inhibitors with the aid of a molecular model of GRK2. We used the available crystal structure of cAMP dependent protein kinase (cAPK) as a template to construct a 3D homology model of GRK2. Known cAPK and GRK2 inhibitors were docked into the active sites of GRK2 and cAPK using DOCK v3.5. H8 docked into the hydrophobic pocket of the adenosine 5'-triphosphate (ATP) binding site of cAPK, consistent with its known competitive cAPK inhibition relative to ATP. Similarly, 3 of 4 known GRK2 inhibitors docked into the ATP binding pocket of GRK2 with good scores. Screening the Fine Chemicals Directory (FCD, containing the 3D structures of 13,000 compounds) for docking into the active sites of GRK2 identified H8 and the known GRK2 inhibitor trifluoperazine as candidates. Whereas H8 indeed inhibited light-dependent phosphorylation of rhodopsin by GRK2, but with low potency, 3 additional FCD compounds with promising GRK2 scores failed to inhibit GRK2. This result demonstrates limitations of the GRK2 model in predicting activity among diverse chemical structures. Docking suramin, an inhibitor of protein kinase C (not present in FCD) yielded a good fit into the ATP binding site of GRK2 over cAPK. Suramin did inhibit GRK2 with IC50 32 μM (pA 2 6.39 for competitive inhibition of ATP). Suramin congeners with fewer sulfonic acid residues (NF062, NF503 [IC50 14 μM]) or representing half of the suramin molecule (NF520) also inhibited GRK2 as predicted by docking. In conclusion, suramin and analogues are lead compounds in the development of more potent and selective inhibitors of GRK2.

An RNA molecule that specifically inhibits G-protein-coupled receptor kinase 2 in vitro

“…Heparin, peptides, and the synthetic compound suramin have been described as potential GRK2 inhibitors (Kim et al 1993;Hasbi et al 2000;Kassack et al 2000;Winstel et al 2005). However, heparin, which inhibits GRK2 at low nanomolar concentrations is highly charged and can only be used when it has direct access to GRKs (Lohse et al 1989).…”

Mechanisms of angiotensin II signaling on cytoskeleton of podocytes

“…Heparin is well known as a potent inhibitor (IC50: 0.15 µM) of G protein-coupled receptor kinase 2 (GRK2). [168][169][170] Furthermore, a recent study determined that ezrin is a novel substrate of GRK2-mediating membrane ruffles in Hep2 cells. 171 Heparin may thus exhibit glomerular protection effects on the podocyte cytoskeleton by inhibiting GRK2-mediated ERM protein phosphorylation.…”

Dynamics of Somatostatin Type 2A Receptor Cargoes in Living Hippocampal Neurons

“…This phenomenon was not observed for ␤-arrestin 2-EGFP. In a first attempt to decipher the intracellular events involved in the translocation of the ␤-arrestin 1 to the nucleus, neurons transfected with sst 2A -WT and ␤-arrestin 1-EGFP were preincubated with pertussis toxin to block Gi/o-mediated signal transduction or with suramin to inhibit the GRK2 (Kassack et al, 2000), a kinase involved in sst 2A receptor phosphorylation and sst 2A receptor-mediated ␤-arrestin mobilization in non-neuronal cells . These pharmacological treatments did not prevent sst 2A receptor internalization and nuclear accumulation of the ␤-arrestin 1 after agonist stimulation (Fig.…”